Using two insulin-sensitive differentiated cell culture systems -the macrophage-like J774.2 and the 3T3-L1 preadipocyte - the state of phosphorylation of the type II cAMP-dependent protein kinase will be assessed under conditions of elevated intracellular cAMP and following treatment with insulin. Correlations will be made of cAMP-dependent protein kinase activity and state of phosphorylation of the cAMP-binding protein. In similar experiments with human lung fibroblasts, the phosphorylation state of the cGMP-dependent protein kinase will be analyzed. The structural defects in the protein kinase mutants of J774.2 will be determined and the synthesis and turnover of wild-type and variant kinases, described. The studies using the macrophage-like cell line will include detection of the cAMP and Ca2+-directed protein phosphorylation reactions involved in macrophage functions using mutants defective in both types of kinases as well as mutants defective in Fc-mediated phagocytosis, motility and plasminogen activator secretion.